Human Antibodies - Volume 12, issue 4

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ISSN 1093-2607 (P)

ISSN 1875-869X (E)

Human Antibodies is an international journal designed to bring together all aspects of human hybridomas and antibody technology under a single, cohesive theme. This includes fundamental research, applied science and clinical applications.

Abstract: Selection from phage antibody libraries can be considered to be an in vitro immune system in which the antibody response is reduced to the bare minimum of antigen recognition. Using selections of antibodies on peptides from a phage antibody library, we investigated what constitutes peptide antigenicity in the context of the antibody-protein binding site. We selected polyclonal antibodies in a high throughput format against 44% of 90 overlapping peptides derived from three different proteins. Of these, 33% of peptides (epitopic peptides) were able to select antibodies that recognized the protein from which the peptides were derived. Although no algorithm was…able to predict all epitopic peptides, solvent accessibility was the best predictor in this cell-free antibody selection context. We subsequently applied solvent accessibility to successfully predict epitopic peptides from p53 and Znf217, and showed that such peptide selected single-chain antibodies were able to recognize soluble p53 in ELISA and Znf217 in a western blot. This is likely to have considerable utility in functional genomics and proteomics where it should be possible to select antibodies against gene products on the basis of deduced amino acid sequence in a high throughput fashion.
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Abstract: Tumor specific peptides recognized by T lymphocytes infiltrating solid tumors, as well as the corresponding T cell receptor (TcR) repertoire usage, have been extensively investigated. By contrast, tumor infiltrating B cells and their immunoglobulin (Ig) repertoire have been studied only in a limited number of tumors. The objective of the present study was to determine, whether DNA sequence analysis of the expressed immunoglobulin variable regions of B cells that infiltrate breast cancer, could be used to reveal a potential specific tumor binding capacity of the antibodies. To answer this question, about 200 expressed Ig heavy (VH) and light chain variable…gene (VL) regions were cloned, sequenced and comparatively analysed from a typical medullary beast carcinoma (MBC), where the massive B and plasma cell infiltration correlates with favourable prognosis despite of its high grade. The tumor infiltrating B cell Ig heavy and light chain sequences could be classified into clusters, families and subgroups, based on the identity level to germline, showing a pattern of oligoclonality. Some overrepresented clusters could be determined. In the course of a detailed analysis and search in Blastn database, a number of VH and VL sequences showed more than 99% homology to DNA sequences of Ig VH region, with proved tumor antigen binding capacity. Our data suggest, that potential tumor binder Ig VH and VL sequences might be selected using a detailed immunoglobulin variable region analysis. This new approach might have a benefit for further antibody engineering, as difficulties in search for tumor binders by phage library selection might be reduced and the time for selection shortened.
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Abstract: The C-erbB-2 proto-oncogene encodes a 185KD glycoprotein with tyrosine kinase activity. Overexpression of this gene either due to gene amplification and/or increased transcription has been observed in a variety of cancers and has been associated with more aggressive disease and a poor clinical prognosis in 20–30% of patients with breast cancer. Besides several prognostic factors like tumor size, histologic grade, steroid hormone receptor status, DNA ploidy, lymph node status etc which are significant in the management of breast cancer, C-erbB-2 status might also serve as an additional parameter. Immunohistochemistry is the most widely used method to study the expression of…C-erbB-2 in breast cancer. The very low levels of expression of C-erbB-2 by normal tissues makes this receptor, a potential target for diagnosis and therapy with monoclonal antibodies raised against its extracellular domain. One such monoclonal antibody designated as CIBCgp185 of IgG2a isotype has been generated in our laboratory and extensively characterized. In the present study, an indirect immunohistochemical assay was carried out on frozen tumor tissue sections of 127 malignant breast tumor specimens of various histological types using monoclonal antibody CIBCgp185, which revealed intense staining of tumor cell membrane in 32 specimens, indicating overexpression of C-erbB-2. In the case of 53 benign breast tissues and 24 normal breast tissues studied, this MAb did not exhibit any reactivity. These results suggest that MAb CIBCgp185 might prove useful to identify tumors with overexpression of C-erbB-2 which are often associated with poor prognosis and early recurrence and might have future therapeutic application in the treatment of these cancers.
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Abstract: Anthrax toxin is the dominant virulence factor of Bacillus anthracis; drugs blocking its action could therefore have therapeutic benefit. We report here the production of a neutralizing monoclonal antibody (mAb) against anthrax lethal factor (LF) and the inhibition by the antibody of anthrax lethal toxin (LeTx) formation. The anti-LF monoclonal antibody LF8 neutralized the LeTx challenge both in vitro with macrophage J774A.1 cells and in vivo in nude mice. Our data suggested that LF8 binds LF at or near the PA binding domain. A set of dodecameric peptides was selected from a phage-displayed peptide library through their specific binding to…anti-LF neutralizing mAb LF8. These small peptides compete with LF to bind LF8. Further investigation is undergoing to test the potential application of these peptides to the clinical treatment of anthrax infection by blocking LeTx formation.
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Abstract: Purpose: SK1, a human IgM monoclonal antibody recognizes the antigen, termed AgSK1 which was shown to be preferentially expressed by human adenocarcinomas, particularly human gastrointestinal malignancies. The aim of this study was to clarify the clinicopathological significance of AgSK1 mRNA expression in human colorectal cancer. Methods: Using a quantitative RT-PCR, we studied the mRNA expression level of AgSK1 in the resected specimen of 40 patients with colorectal cancer. We estimated the tumor tissue value (T-value), nontumorous tissue value (N-value), and the ratio of T-value to N-value (T/N Ratio). For further analysis, we compared these data with the clinicopathological…features. Results: In the status of lymph node metastases, pN positive status tended to have a higher T-value level of AgSK1 mRNA than pN negative status (P = 0.076). According to a mean T-value (8.032) or a mean T/N Ratio (1.549), we divided these patients into two groups, low expression group and high expression group. A high expression group showed a significantly higher frequency of positive lymph node metastases (T-value; P = 0.021, T/N Ratio; P = 0.024). Conclusion: AgSK1 mRNA expression in tumor tissue may become a useful marker for lymph node metastases and a malignant potential marker of colorectal cancer.
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